Role of PDGFRA+ cells and a CD55+ PDGFRALo fraction in the gastric mesenchymal niche

PDGFRA-expressing mesenchyme supports intestinal stem cells. Stomach epithelia have related niche dependencies, but their enabling mesenchymal cell populations are unknown, in part because previous studies pooled the gastric antrum and corpus. Our high-resolution imaging, transcriptional profiling, and organoid assays identify regional subpopulations and supportive capacities of purified mouse corpus and antral PDGFRA+ cells. Sub-epithelial PDGFRAHi myofibroblasts are principal sources of BMP ligands and two molecularly distinct pools distribute asymmetrically along antral glands but together fail to support epithelial growth in vitro. In contrast, PDGFRALo CD55+ cells strategically positioned beneath gastric glands promote epithelial expansion in the absence of other cells or factors. This population encompasses a small fraction expressing the BMP antagonist Grem1. Although Grem1+ cell ablation in vivo impairs intestinal stem cells, gastric stem cells are spared, implying that CD55+ cell activity in epithelial self-renewal derives from other subpopulations. Our findings shed light on spatial, molecular, and functional organization of gastric mesenchyme and the spectrum of signaling sources for epithelial support.

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-Images were analyzed using Fiji and ZEN Desk 3.4 software.
-Videos were generated with ZEN Desk 3.4 software.
-GFP+ cell images were quantified using Cellpose 2.0 plugin in in CellProfiler 4.2.1 -GFP signal was quantified using Cellpose 2.0 and feeding segmented masks to to CellProfiler 4.2.1 -GraphPad Prism version 9 -R platform 4.2.1 -Bulk RNAseq analysis: Data were aligned to to mouse reference genome mm10 using the Viper pipeline with default settings.Data quality was verified using RSeQC.Data were normalized and differential gene expression (padj <0.05; |log2 fold-change| >1.5) was analyzed with the DESeq2 package.Pearson correlation coefficients were calculated from DESeq2 normalized counts and plotted using Corrplot package.Integrative Genome Viewer (IGV) data tracks were generated from RPKM-normalized bigwigs loaded into IGV 2.15.4 (Broad Institute).
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No statistical methods were used to determine sample size -Reads from genes known to express in gastric epithelium and genes encoded on sex chromosomes were excluded from Bulk RNAseq analysis -For scRNAseq analysis, cells were filtered for <12% mitochondrial and >1,000 unique reads.
instrument (Roche) for quantitative PCR analysis -Cells were sorted on on a BD BD FACSAria II II cell sorter with BD BD FACSDiva 8.0.1 software -Data from cells were collected by by FACS on on a BD BD LSR Fortessa cell analyzer -Images were taken using an an SP5X laser scanning confocal microscope (Leica Microsystems CMS GmbH) or or an an LSM980 microscope and ZEN Desk 3.4 software (Carl Zeiss GmbH).
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We require information from authors about some types of materials, experimental systems and methods used in many studies.Here, indicate whether each material, system or method listed is relevant to your study.If you are not sure if a list item applies to your research, read the appropriate section before selecting a response.Biological replicates were performed at least 3 times.All attempts at replication were successful.The mean +/-S.E.M, and statistical analysis (ANOVA Tukey's multiple comparison test, one-way ANOVA Dunnett's multiple comparison test, unpaired two-tailed t-test) was used to see if and how the difference between parameters are significant.Significance were presented as stars and p values, calculated by Graph Pad software.Samples were not randomized for this studyThere was no blinded allocation of samples during experiments or analysis nature portfolio | reporting summary